BACKGROUND Gram-negative enteric rods were associated with periodontal diseases in several populations. The association between enteric and putative periodontal pathogens has received little attention in the literature. The present investigation determined the occurrence and in vitro antimicrobial susceptibility of isolates of Gram-negative enteric rods from Colombian patients with chronic periodontitis. METHODS Clinical parameters and the in vitro antimicrobial sensitivity of isolates of Gram-negative enteric rods were examined in 76 patients with chronic periodontitis in Medellin, Colombia. Subgingival samples were processed using culture and biochemical tests for the detection of periodontal and superinfecting pathogens. Selected colonies of Gram-negative enteric rods from pure cultures were used to test the susceptibility to amoxicillin/clavulanic acid, ciprofloxacin, and moxifloxacin. Chi-square and Mann-Whitney tests were used to determine differences in clinical variables versus the presence or absence of Gram-negative enteric rods. RESULTS Klebsiella pneumoniae was found in 12 patients, Pseudomonas aeruginosa was found in four patients, and three other species were recovered with a lower prevalence. Men (38.75%) tended to harbor more of the studied organisms than women (17.7%) (P = 0.04). Gram-negative enteric rods in periodontal pockets correlated positively with the presence of Aggregatibacter actinomycetemcomitans (previously Actinobacillus actinomycetemcomitans), Porphyromonas gingivalis, and Prevotella intermedia/nigrescens (respectively, r = 0.66, 0.31, and 0.32; P <0.001). All superinfecting organisms demonstrated a high susceptibility to moxifloxacin and ciprofloxacin but exhibited a variable susceptibility to amoxicillin/clavulanic acid. CONCLUSIONS This study found high levels of Gram-negative enteric rods in patients with chronic periodontitis. Moxifloxacin and ciprofloxacin appeared capable of eradicating these organisms from periodontal pockets. Its good activity against Gram-negative enteric rods and periodontopathogens suggests the potential use of moxifloxacin as an adjunctive antibiotic in the treatment of mixed periodontal infections.